Arctiin Suppresses H9N2 Avian Influenza Virus-Mediated Inflammation Via Activation of Nrf2/HO-1 Signaling

Abstract Background: H9N2 avian influenza viruses (AIVs) infect avian and mammalian hosts and provide internal genes for new emerging highly pathogenic avian viruses that cause severe pneumonia with high mortality, for which few medications are available. Arctiin, a bioactive lignan glycoside, has been reported to possess multiple pharmacological properties. However, the effect of arctiin on H9N2 virus infection is unclear. In the current study, we analyzed the effect of arctiin on H9N2 virus infection and the underlying molecular mechanism in vitro. Methods: The antiviral effect against H9N2 virus was determined by plaque reduction assay (PRA) and progeny virus reduction assay. We employed MTT assay, qRT-PCR, ELISA, immunofluorescence and Western blotting to better understand the anti-inflammatory effect and corresponding mechanism of arctiin on H9N2 virus-infected cells.Results: The results showed that arctiin had antiviral activity against H9N2 virus. Arctiin treatment reduced H9N2 virus-triggered proinflammatory cytokines, such as IL-6, and TNF-α. Moreover, arctiin significantly suppressed H9N2 virus-mediated expression of COX-2 and PGE2. Furthermore, we found that arctiin inhibited H9N2 virus-mediated activation of RIG-I/JNK MAPK signaling. Interestingly, arctiin treatment obviously reversed H9N2 virus-induced reduction of Nrf2, increased the nuclear translocation of Nrf2, and upregulated Nrf2 signaling target genes (HO-1 and SOD2). Zinc protoporphyrin (Znpp)—an HO-1 inhibitor—weakened the inhibitory effect of arctiin on H9N2 virus-induced RIG-I/JNK MAPK and proinflammatory mediators. Conclusion: Taken together, these results suggested that the anti-inflammatory effects of arctiin on H9N2 virus infection may be due to the activation of Nrf2/HO-1 and blocked RIG-I/JNK MAPK signaling; thus, arctiin may be a promising agent for prevention and treatment of H9N2 virus infections.

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Arctiin suppresses H9N2 avian influenza virus-mediated inflammation via activation of Nrf2/HO-1 signaling

BMC Complementary Medicine and Therapies ◽

10.1186/s12906-021-03462-4 ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Beixian Zhou ◽

Linxin Wang ◽

Yueyun Liang ◽

Jing Li ◽

Xiping Pan

Keyword(s):

Virus Infection ◽

Avian Influenza ◽

Nuclear Translocation ◽

Mapk Signaling ◽

Influenza Viruses ◽

H9n2 Virus ◽

Progeny Virus ◽

Zinc Protoporphyrin ◽

Reduction Assay ◽

Anti Inflammatory

Abstract Background H9N2 avian influenza viruses (AIVs) infect avian and mammalian hosts and provide internal genes for new emerging highly pathogenic avian viruses that cause severe pneumonia with high mortality, for which few medications are available. Arctiin, a bioactive lignan glycoside, has been reported to possess multiple pharmacological properties. However, the effect of arctiin on H9N2 virus infection is unclear. In the current study, we analyzed the effect of arctiin on H9N2 virus infection and the underlying molecular mechanism in vitro. Methods The antiviral effect against H9N2 virus was determined by plaque reduction assay (PRA) and progeny virus reduction assay. We employed MTT assay, qRT-PCR, ELISA, immunofluorescence and Western blotting to better understand the anti-inflammatory effect and corresponding mechanism of arctiin on H9N2 virus-infected cells. Results The results showed that arctiin had antiviral activity against H9N2 virus. Arctiin treatment reduced H9N2 virus-triggered proinflammatory cytokines, such as IL-6, and TNF-α. Moreover, arctiin significantly suppressed H9N2 virus-mediated expression of COX-2 and PGE2. Furthermore, we found that arctiin inhibited H9N2 virus-mediated activation of RIG-I/JNK MAPK signaling. Interestingly, arctiin treatment obviously reversed H9N2 virus-induced reduction of Nrf2, increased the nuclear translocation of Nrf2, and upregulated Nrf2 signaling target genes (HO-1 and SOD2). Zinc protoporphyrin (Znpp)—an HO-1 inhibitor—weakened the inhibitory effect of arctiin on H9N2 virus-induced RIG-I/JNK MAPK and proinflammatory mediators. Conclusion Taken together, these results suggested that the anti-inflammatory effects of arctiin on H9N2 virus infection may be due to the activation of Nrf2/HO-1 and blocked RIG-I/JNK MAPK signaling; thus, arctiin may be a promising agent for prevention and treatment of H9N2 virus infections.

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Anti-Inflammatory Activity of Populus deltoides Leaf Extract via Modulating NF-κB and p38/JNK Pathways

International Journal of Molecular Sciences ◽

10.3390/ijms19123746 ◽

2018 ◽

Vol 19 (12) ◽

pp. 3746 ◽

Cited By ~ 5

Author(s):

Ye Jeong ◽

Mi-Young Lee

Keyword(s):

Nitric Oxide ◽

Protein Kinase ◽

Leaf Extract ◽

Populus Deltoides ◽

Nuclear Translocation ◽

Mapk Signaling ◽

Mitogen Activated Protein Kinase ◽

Inflammatory Activity ◽

No Production ◽

Anti Inflammatory

Populus deltoides, known as eastern cottonwood, has been commonly used as a medicinal plant. The aim of the present study was to investigate the mechanism underlying the anti-inflammatory activity of P. deltoides leaf extract (PLE). PLE effectively inhibited the expression of inducible nitric oxide synthase (iNOS) and nitric oxide (NO) production in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells, but not that of cyclooxygenase-2 (COX-2) and prostaglandin E2. Proinflammatory tumor necrosis factor alpha (TNF-α) levels were also reduced by the extract. PLE inhibited the phosphorylation of nuclear factor-kappa B (NF-κB) and inhibitor of Kappa Bα (IκBα), and blunted LPS-triggered enhanced nuclear translocation of NF-κB p65. In mitogen-activated protein kinase (MAPK) signaling, PLE effectively decreased the phosphorylation of p38 and c-Jun N-terminal protein kinase (JNK), but not of extracellular signal-regulated kinase 1/2 (ERK1/2). Taken together, these results suggest that anti-inflammatory activity of P. deltoides leaf extract might be driven by iNOS and NO inhibition mediated by modulation of the NF-κB and p38/JNK signaling pathways.

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The PB2 and M genes are critical for the superiority of genotype S H9N2 virus to genotype H in optimizing viral fitness of H5Nx and H7N9 avian influenza viruses in mice

Transboundary and Emerging Diseases ◽

10.1111/tbed.13395 ◽

2019 ◽

Vol 67 (2) ◽

pp. 758-768

Author(s):

Xiaoli Hao ◽

Jiao Hu ◽

Xiaoquan Wang ◽

Min Gu ◽

Jiongjiong Wang ◽

...

Keyword(s):

Avian Influenza ◽

Influenza Viruses ◽

Viral Fitness ◽

H9n2 Virus ◽

Avian Influenza Viruses

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Cerevisterol Alleviates Inflammation via Suppression of MAPK/NF-κB/AP-1 and Activation of the Nrf2/HO-1 Signaling Cascade

Biomolecules ◽

10.3390/biom10020199 ◽

2020 ◽

Vol 10 (2) ◽

pp. 199 ◽

Cited By ~ 2

Author(s):

Md Badrul Alam ◽

Nargis Sultana Chowdhury ◽

Md Hossain Sohrab ◽

Md Sohel Rana ◽

Choudhury Mahmood Hasan ◽

...

Keyword(s):

Endophytic Fungi ◽

Fusarium Solani ◽

Nuclear Translocation ◽

Mapk Signaling ◽

Inflammatory Activity ◽

Signaling Cascade ◽

Nuclear Factor ◽

Tnf Α ◽

Gene Assay ◽

Anti Inflammatory

As part of our continuous effort to find potential anti-inflammatory agents from endophytic fungi, a Fusarium solani strain, isolated from the plant Aponogeton undulatus Roxb., was investigated. Cerevisterol (CRVS) was identified from endophytic fungi, a Fusarium solani strain, and moreover exhibited anti-inflammatory activity. However, the underlying mode of action remains poorly understood. The aim of this study is to reveal the potential mechanisms of CRVS against inflammation on a molecular level in LPS-activated RAW 264.7 peritoneal macrophage cells. CRVS was isolated from F. solani and characterized based on spectral data analysis. The MTT assay was performed to measure cell viability in CRVS-treated macrophages. Anti-inflammatory activity was assessed by measurement of nitric oxide (NO) and prostaglandin E2 (PGE2) levels, as well as the production of various cytokines, such as tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and -6 (IL-6) in LPS-stimulated macrophages. RT-PCR and immunoblotting analyses were done to examine the expression of various inflammatory response genes. A reporter gene assay was conducted to measure the level of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and activator protein-1 (AP-1) transactivation. CRVS suppresses the LPS-induced production of NO and PGE2, which is a plausible mechanism for this effect is by reducing the expression of iNOS and COX-2. CRVS also decreases the expression of pro-inflammatory cytokines, such as TNF-α, IL-6, and IL-1β. CRVS halted the nuclear translocation of NF-κB by blocking the phosphorylation of inhibitory protein κBα (IκBα) and suppressing NF-κB transactivation. The mitogen-activated protein kinases (MAPK) signaling pathways are also suppressed. CRVS treatment also inhibited the transactivation of AP-1 and the phosphorylation of c-Fos. Furthermore, CRVS could induce the nuclear translocation of nuclear factor erythroid 2-related factor 2 (Nrf2) by down-regulating Kelch-like ECH-associated protein 1 (Keap-1) and up-regulating hemeoxygenases-1 (HO-1) expression. The results suggest that CRVS acts as a natural agent for treating inflammatory diseases by targeting an MAPK, NF-κB, AP-1, and Nrf2-mediated HO-1 signaling cascade.

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Tropism and Infectivity of a Seasonal A(H1N1) and a Highly Pathogenic Avian A(H5N1) Influenza Virus in Primary Differentiated Ferret Nasal Epithelial Cell Cultures

Journal of Virology ◽

10.1128/jvi.00080-19 ◽

2019 ◽

Vol 93 (10) ◽

Cited By ~ 6

Author(s):

Hui Zeng ◽

Cynthia S. Goldsmith ◽

Amrita Kumar ◽

Jessica A. Belser ◽

Xiangjie Sun ◽

...

Keyword(s):

Influenza Virus ◽

Virus Infection ◽

Avian Influenza ◽

H5n1 Virus ◽

H1n1 Virus ◽

Influenza Virus Infection ◽

Influenza Viruses ◽

Host Interactions ◽

Culture Model

ABSTRACTFerrets represent an invaluable animal model to study influenza virus pathogenesis and transmission. To further characterize this model, we developed a differentiated primary ferret nasal epithelial cell (FNEC) culture model for investigation of influenza A virus infection and virus-host interactions. This well-differentiated culture consists of various cell types, a mucociliary clearance system, and tight junctions, representing the nasal ciliated pseudostratified respiratory epithelium. Both α2,6-linked and α2,3-linked sialic acid (SA) receptors, which preferentially bind the hemagglutinin (HA) of human and avian influenza viruses, respectively, were detected on the apical surface of the culture with different cellular tropisms. In accordance with the distribution of SA receptors, we observed that a pre-2009 seasonal A(H1N1) virus infected both ciliated and nonciliated cells, whereas a highly pathogenic avian influenza (HPAI) A(H5N1) virus primarily infected nonciliated cells. Transmission electron microscopy revealed that virions were released from or associated with the apical membranes of ciliated, nonciliated, and mucin-secretory goblet cells. Upon infection, the HPAI A(H5N1) virus replicated to titers higher than those of the human A(H1N1) virus at 37°C; however, replication of the A(H5N1) virus was significantly attenuated at 33°C. Furthermore, we found that infection with the A(H5N1) virus induced higher expression levels of immune mediator genes and resulted in more cell damage/loss than with the human A(H1N1) virus. This primary differentiated FNEC culture model, recapitulating the structure of the nasal epithelium, provides a useful model to bridgein vivoandin vitrostudies of cellular tropism, infectivity, and pathogenesis of influenza viruses during the initial stages of infection.IMPORTANCEAlthough ferrets serve as an important model of influenza virus infection, much remains unknown about virus-host interactions in this species at the cellular level. The development of differentiated primary cultures of ferret nasal epithelial cells is an important step toward understanding cellular tropism and the mechanisms of influenza virus infection and replication in the airway milieu of this model. Using lectin staining and microscopy techniques, we characterized the sialic acid receptor distribution and the cellular composition of the culture model. We then evaluated the replication of and immune response to human and avian influenza viruses at relevant physiological temperatures. Our findings offer significant insight into this first line of defense against influenza virus infection and provide a model for the evaluation of emerging influenza viruses in a well-controlledin vitroenvironmental setting.

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Constitutively Expressed IFITM3 Protein in Human Endothelial Cells Poses an Early Infection Block to Human Influenza Viruses

Journal of Virology ◽

10.1128/jvi.01254-16 ◽

2016 ◽

Vol 90 (24) ◽

pp. 11157-11167 ◽

Cited By ~ 15

Author(s):

Xiangjie Sun ◽

Hui Zeng ◽

Amrita Kumar ◽

Jessica A. Belser ◽

Taronna R. Maines ◽

...

Keyword(s):

Endothelial Cells ◽

Influenza Virus ◽

Virus Infection ◽

Avian Influenza ◽

Influenza Virus Infection ◽

Influenza Viruses ◽

Human Influenza ◽

Avian Influenza Viruses ◽

Human Influenza Virus ◽

Pulmonary Endothelial Cells

ABSTRACTA role for pulmonary endothelial cells in the orchestration of cytokine production and leukocyte recruitment during influenza virus infection, leading to severe lung damage, has been recently identified. As the mechanistic pathway for this ability is not fully known, we extended previous studies on influenza virus tropism in cultured human pulmonary endothelial cells. We found that a subset of avian influenza viruses, including potentially pandemic H5N1, H7N9, and H9N2 viruses, could infect human pulmonary endothelial cells (HULEC) with high efficiency compared to human H1N1 or H3N2 viruses. In HULEC, human influenza viruses were capable of binding to host cellular receptors, becoming internalized and initiating hemifusion but failing to uncoat the viral nucleocapsid and to replicate in host nuclei. Unlike numerous cell types, including epithelial cells, we found that pulmonary endothelial cells constitutively express a high level of the restriction protein IFITM3 in endosomal compartments. IFITM3 knockdown by small interfering RNA (siRNA) could partially rescue H1N1 virus infection in HULEC, suggesting IFITM3 proteins were involved in blocking human influenza virus infection in endothelial cells. In contrast, selected avian influenza viruses were able to escape IFITM3 restriction in endothelial cells, possibly by fusing in early endosomes at higher pH or by other, unknown mechanisms. Collectively, our study demonstrates that the human pulmonary endothelium possesses intrinsic immunity to human influenza viruses, in part due to the constitutive expression of IFITM3 proteins. Notably, certain avian influenza viruses have evolved to escape this restriction, possibly contributing to virus-induced pneumonia and severe lung disease in humans.IMPORTANCEAvian influenza viruses, including H5N1 and H7N9, have been associated with severe respiratory disease and fatal outcomes in humans. Although acute respiratory distress syndrome (ARDS) and progressive pulmonary endothelial damage are known to be present during severe human infections, the role of pulmonary endothelial cells in the pathogenesis of avian influenza virus infections is largely unknown. By comparing human seasonal influenza strains to avian influenza viruses, we provide greater insight into the interaction of influenza virus with human pulmonary endothelial cells. We show that human influenza virus infection is blocked during the early stages of virus entry, which is likely due to the relatively high expression of the host antiviral factors IFITMs (interferon-induced transmembrane proteins) located in membrane-bound compartments inside cells. Overall, this study provides a mechanism by which human endothelial cells limit replication of human influenza virus strains, whereas avian influenza viruses overcome these restriction factors in this cell type.

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Molecular immunophenotyping of lungs in naïve and vaccinated chickens early after pulmonary avian influenza A (H9N2) virus infection

Veterinary Immunology and Immunopathology ◽

10.1016/j.vetimm.2008.10.245 ◽

2009 ◽

Vol 128 (1-3) ◽

pp. 325

Author(s):

Winfried G.J. Degen ◽

Jacqueline Smith ◽

Bartjan Simmelink ◽

Elizabeth J. Glass ◽

Dave W. Burt ◽

...

Keyword(s):

Virus Infection ◽

Avian Influenza ◽

Influenza A ◽

H9n2 Virus

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Contribution of Segment 3 to the Acquisition of Virulence in Contemporary H9N2 Avian Influenza Viruses

Journal of Virology ◽

10.1128/jvi.01173-20 ◽

2020 ◽

Vol 94 (20) ◽

Author(s):

Anabel L. Clements ◽

Joshua E. Sealy ◽

Thomas P. Peacock ◽

Jean-Remy Sadeyen ◽

Saira Hussain ◽

...

Keyword(s):

Middle East ◽

Avian Influenza ◽

Mortality Rates ◽

Influenza Viruses ◽

H9n2 Virus ◽

Morbidity And Mortality ◽

Single Amino Acid ◽

Economic Damage ◽

Poultry Production ◽

Avian Influenza Viruses

ABSTRACT H9N2 avian influenza viruses (AIVs) circulate in poultry throughout much of Asia, the Middle East, and Africa. These viruses cause huge economic damage to poultry production systems and pose a zoonotic threat both in their own right and in the generation of novel zoonotic viruses, for example, H7N9. In recent years, it has been observed that H9N2 viruses have further adapted to gallinaceous poultry, becoming more highly transmissible and causing higher morbidity and mortality. Here, we investigate the molecular basis for this increased virulence, comparing a virus from the 1990s and a contemporary field strain. The modern virus replicated to higher titers in various systems, and this difference mapped to a single amino acid polymorphism at position 26 of the endonuclease domain shared by the PA and PA-X proteins. This change was responsible for increased replication and higher morbidity and mortality rates along with extended tissue tropism seen in chickens. Although the PA K26E change correlated with increased host cell shutoff activity of the PA-X protein in vitro, it could not be overridden by frameshift site mutations that block PA-X expression and therefore increased PA-X activity could not explain the differences in replication phenotype. Instead, this indicates that these differences are due to subtle effects on PA function. This work gives insight into the ongoing evolution and poultry adaptation of H9N2 and other avian influenza viruses and helps us understand the striking morbidity and mortality rates in the field, as well as the rapidly expanding geographical range seen in these viruses. IMPORTANCE Avian influenza viruses, such as H9N2, cause huge economic damage to poultry production worldwide and are additionally considered potential pandemic threats. Understanding how these viruses evolve in their natural hosts is key to effective control strategies. In the Middle East and South Asia, an older H9N2 virus strain has been replaced by a new reassortant strain with greater fitness. Here, we take representative viruses and investigate the genetic basis for this “fitness.” A single mutation in the virus was responsible for greater fitness, enabling high growth of the contemporary H9N2 virus in cells, as well as in chickens. The genetic mutation that modulates this change is within the viral PA protein, a part of the virus polymerase gene that contributes to viral replication as well as to virus accessory functions—however, we find that the fitness effect is specifically due to changes in the protein polymerase activity.

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